Application of k-carrageenan oligosaccharides capable of adjusting activated states of gitter cell
A technology of microglia and carrageenan oligosaccharides, applied in the field of κ-carrageenan oligosaccharides in medicine and health food, can solve the problems of aggravating nerve cell damage and achieve the effect of improving the body's immunity
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Embodiment 1
[0023] Example 1: Effect of κ-carrageenan oligosaccharide (KCOS) on the proliferation of microglial cells
[0024] 1.1 Inverted microscope observation results of the effect of κ-carrageenan oligosaccharides on the proliferation of microglial cells
[0025] Cell culture conditions: IMDM (sigma, USA) medium containing 10% calf serum, 100 μg / ml streptomycin and 100IU / ml penicillin, 5% CO 2 , 37°C, subculture every two days. The day before the experiment, the N9 medium was changed. Wash 3 times with PBS, digest with 0.125% trypsin, pipette, and make cell density 5×10 with IMDM medium containing 10% fetal bovine serum 4 cells / mL of cell suspension. In a 96-well culture plate, add 200 μl of cell suspension to each well. After culturing for 12 hours, carefully absorb the culture medium and wash it with PBS. In the experimental group, add different concentrations of KCOS and DKCOS (the desulfurization group of κ-carrageenan oligosaccharide) to each well. derivatives), so that the ...
Embodiment 2
[0030] Example 2: Effects of κ-carrageenan oligosaccharides on cytokines secreted by microglia
[0031] 2.1 Griess method test results of the effect of κ-carrageenan oligosaccharides on the release of NO from microglia
[0032] NO is easily oxidized to NO in the body or in aqueous solution 2- , under acidic conditions, NO 2- The diazonium reaction occurs with the diazonium salt sulfonamide, and a diazo compound is generated, and the latter further undergoes a coupling reaction with naphthyl vinyl diamine, and the concentration of the product generated by the reaction is the same as that of NO 2- The concentration has a linear relationship, and there is a maximum absorption peak at 540nm. Determination of NO / NO by Griess method 2- Take the cell culture supernatant and add an equal volume of Griess reagent (1.0% anhydrous sulfanilic acid, 0.10% N-1-naphthylethylenediamine hydrochloride, 2.5% phosphoric acid), and measure the absorbance at 540 nm.
[0033] Results: NO is an i...
Embodiment 3
[0037] Example 3: The way κ-carrageenan oligosaccharides act on microglia
[0038] 3.1 Laser confocal detection of the effect of FITC-labeled κ-carrageenan oligosaccharides on microglia
[0039] The pure product of fluorescein isothiocyanate (FITC) is yellow or orange-yellow crystalline powder, easily soluble in water and alcohol solvents. The molecular weight is 389.4, the maximum absorption wavelength is 490-495nm, and the maximum emission wavelength is 520-530nm. It presents bright yellow-green fluorescence and is currently the most widely used fluorescein. Since κ-carrageenan oligosaccharide itself has a negative charge, it can be combined with FITC through charge interaction, and can be used to indicate the interaction between oligosaccharides and cells under a fluorescence microscope.
[0040] 3.1.1 FITC-labeled κ-carrageenan oligosaccharides:
[0041] Mix κ-carrageenan oligosaccharide with equal mass of FITC, dissolve in a certain amount of distilled water, stir at ro...
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